Immunological and biochemical analyses revealed that C-ASWS a mycelial-phase cell wall antigen of Coccidioides immitis contained 1% amino sugar, 85% neutral sugars, and 10% protein. The neutral sugar composition was mannose, galactose, glucose, and 3-0-methylmannose Two-dimensional immunoelectrophoresis using antiserum from a burro hyperimmunized with coccidioidin revealed that C-ASWS contained 8 protein-containing components. All 8 bands were in common with coccidioidin (total of 26 bands); 7 were in common with spherulin (12 bands). Comparative studies showed that 5 protein-staining bands were detectable in histoplasmin H42 with the anti-coccidioidin serum. Four of these 5 bands corresponded to 4 bands in C-ASWS and in coccidioidin and spherulin. As would be expected, a high degree of cross reactivity was detected with C-ASWS, coccidioidin, and spherulin in in vitro lymphocyte transformation assays of healthy histoplasmin skin-test positive persons. In contrast, only minimal cross reactivity was obtained with histoplasmin using lymphocytes from coccidioidin skin-test positive subjects. Both C-ASWS and coccidioidin were equally effective in eliciting MIF responses in C-ASWS sensitized guinea pigs. Spherulin failed to elicit significant MIF responses. No cross reactivity was detected when histoplasmin was assayed using macrophages from these same guinea pigs. Studies are now in progress to isolate the individual protein-containing bands in each of the 3 Coccidioides antigens and in histoplasmin and to assay the biological activity of the various components.